CD40L (CD154;gp39) is a critical effector molecule of activated T cells, necessary for production of isotype switched and affinity matured antibodies by B cells, and macrophage and dendritic cell activation. Abnormal CD40L expression by T and B cells is thought to drive multiple human immunological and inflammatory diseases of which systemic lupus erythematosus (Lupus, SLE) and rheumatoid arthritis are most prominent. Consequently, identification of factors that regulate CD40L is necessary to better understand the etiology of these diseases and possibly develop new treatment strategies. Studies from the applicant now show that the related transcription factors TFE3 and TFEB are physiological and direct activators of CD40L gene expression in T cells. Simultaneous inactivation of TFE3 and TFEB exclusively in T cells in mice resulted in an immune deficiency resembling human Hyper IgM syndrome caused by CD40L deficiency, characterized by defective humoral immune responses to thymus (T)-dependent antigens, poor germinal center formation, but normal T-independent humoral responses. T cells from such mice exhibited impaired CD40L expression. This discovery was possible by expressing a transdominant-negative (TON) inhibitory protein that simultaneously blocked TFE3 and TFEB activity in T cells via transgenesis. This was necessary because genetic TFEB-deficiency causes early embryonic death and TFE3 and TFEB are functionally redundant with respect to CD40L. The purpose of the proposed studies is to further define how TFE3 and TFEB contribute to immune function and the autoimmune disease SLE primarily via their role in governing CD40L expression in lymphocytes. Experiments in Aim 1 will molecularly define the conditions and means by which TFE3 and TFEB control CD40L expression in response to T cell stimulation in mouse and human T cells. In Aim 2, we will evaluate the contribution of TFE3- and TFEB-dependent CD40L expression to the development of autoimmune disease in the Lupus-prone MRL/lpr mouse. In Aim 3, collaborative studies with Dr. E. Ginzler, who runs the SUNY-Downstate Lupus Cohort, will evaluate the status and contribution of TFE3 and TFEB to abnormal CD40L expression in lymphocytes from patients with SLE. Such information is important to fully understand the molecular basis of this immune pathology, to devise new treatment strategies, and possibly identify new prognostic markers.